A tissue engineered pancreatic substitute has the potential to provide a less invasive, more physiologic and potentially less costly regulation of blood glucose levels than current treatment methods based on insulin injections or insulin pumps. However, the development of tissue based therapies is hampered by the low availability of donor tissue and/or the need to immunosuppress the transplant recipient. Tissue substitutes based on non-pancreatic cells retrieved from the same patient and engineered for physiologically responsive insulin secretion have the potential to overcome both of these limitations. The long-range goal associated with this research program is to produce the fundamental knowledge and enabling technologies for developing an efficacious and immune acceptable tissue substitute based on such cells. The objective of this application is to engineer a pancreatic substitute consisting of two components: one, based on recombinant hepatic cells and the second, based on recombinant enteroendocrine cells. The central hypothesis is that these two components, in concert, exhibit insulin secretion dynamics closely approximating those of pancreatic islets and can restore normoglycemia more effectively than either component alone. Guided by strong preliminary data, the hypothesis will be addressed by the following three specific aims: 1) develop a pancreatic substitute based on recombinant hepatocytes secreting insulin under transcriptional regulation;2) develop a substitute based on recombinant, insulin-secreting intestinal endocrine cells, which exhibit physiologic responsiveness at the secretion pathway level;3) combine the constructs developed in Aims 1 and 2 to generate a substitute that exhibits appropriate insulin secretion dynamics for in vivo efficacy. The approach is innovative, as it engineers a functional tissue by combining two cell types with each providing a component of the desired functionality. The proposed research is significant, as it will generate new fundamental information on the development of autologous cell therapies for diabetes, which are not limited by tissue availability and immunoreactivity. It is expected that the generated knowledge will produce a significant advancement towards a therapy that is applicable at a clinically relevant scale.